1. Field of the Invention
The present invention relates to a method for separating .alpha.-L-aspartyl-L-phenylalanine methyl ester (.alpha.-APM) from a solution containing .alpha.-APM and impurities associated with the production thereof.
2. Description of the Prior Art
.alpha.-L-aspartyl-L-phenylalanine methyl ester (hereinafter abbreviated as .alpha.-APM) has been widely known as a dipeptide base sweetener. It has sweetness of good quality and a sweetness degree of about 200 times the sweetness of sucrose. The demand for .alpha.-APM is now rapidly expanding as a diet sweetener.
.alpha.-APM is a dipeptide compound composed of L-aspartic acid and L-phenylalanine methyl ester. Various methods have already been disclosed for the preparation of .alpha.-APM. These methods are primarily chemical processes and generally employ L-aspartic acid anhydride having a protected amino group as a starting material.
For example, it has been known that L-aspartic acid anhydride having a protected amino group can be subjected to a condensation reaction with L-phenylalanine methyl ester in a suitable solvent, and the protective group can be subsequently cleaved with a usual method to obtain .alpha.-APM (U.S. Pat. No. 3,786,039). A process for the preparation of .alpha.-APM from materials other than L-phenylalanine methyl ester is disclosed in U.S. Pat. No. 3,933,781. In the process, N-formyl-L-aspartic acid anhydride is subjected to a condensation reaction with L-phenylalanine in acetic acid and subsequently deformylated in the presence of hydrogen halide. The resultant intermediate is esterified by treating with water, alcohol and hydrogen halide. Then .alpha.-APM is isolated in the form of a hydrogen halide salt.
U.S. Pat. No. 4,173,562 describes another process in which N-formyl-L-aspartic acid anhydride is subjected to a condensation reaction with L-phenylalanine, and then deformylation and esterification are simultaneously carried out to produce .alpha.-APM.
U.S Pat. No. 3,962,207 also teaches a process using a mineral acid salt of L-aspartic acid anhydride. The mineral acid salt is reacted with L-phenylalanine methyl ester in a solvent mixture consisting of strong acid, water and methanol. .alpha.-APM is isolated from the solvent mixture in the form of a strong acid salt of .alpha.-APM.
In each of the above processes, however, .beta.-L-aspartyl-L-phenylalanine methyl ester (hereinafter abbreviated as .beta.-APM) is inevitably formed as a by-product in addition to the desired .alpha.-APM. .beta.-APM has no sweet flavor and instead exhibits a bitter taste. Therefore, contamination therewith decreases the commodity value of .alpha.-APM.
When an N-acyl group, for example, an N-formyl group, is used as a protective group, the protective group is generally removed by bringing the group into contact with a strong acid.
Under these conditions, however, the methyl ester group of .alpha.-APM is liable to undergo partial hydrolysis. In the presence of methanol, .alpha.-L-aspartyl-L-phenylalanine-.beta.-methyl ester and/or .alpha.-L-aspartyl-L-phenylalanine dimethyl ester are formed as by-products, and thus it is difficult to selectively obtain only the desired .alpha.-APM.
On the other hand, when a benzyloxycarbonyl group is used as the protective group, the protective group can be readily removed by catalytic reduction, thereby eliminating the problem of hydrolyzing methyl ester. Thus it is known that this protective group is capable of being removed with high selectivity. Japanese Patent Publication Nos. 25537/1982 and 25538/1982 disclose a process wherein benzyloxycarbonylaspartic acid anhydride is reacted with L-phenylalanine methyl ester, the resultant N-benzyloxycarbonyl-L-aspartyl-L-phenylalanine methyl ester (hereinafter abbreviated as Z-APM) is hydrogenated in the presence of an aqueous mineral acid solution, and the reaction mixture thus obtained is neutralized to give .alpha.-APM. The process, however, also forms .beta.-APM as a by-product and an aqueous mineral acid solution is employed in order to remove the protective group. Therefore, the methyl, ester group of .alpha.-APM is apt to hydrolyze while removing the N-benzyloxycarbonyl group. A catalytic reduction process in an acetic acid is also described in the patent publication. Diketopiperazine derivatives are formed in the step of distilling off acetic acid after completing the catalytic reduction, which decreases the yield of .alpha.-APM. The diketopiperazine derivatives also have no sweet flavor and their contamination adversely affects the commodity value of .alpha.-APM.
In addition, it is known as a conventional separation method to bring a mixture of .alpha.- and .beta.-APM into contact with .beta.-resorcylic acid in an aqueous medium. In the method, .alpha.-APM forms a slightly soluble adduct and separates from the contaminating .beta.-APM (Japanese Patent Publication No. 6305/1974).
Although the method can separate .alpha.-APM from its impurities contained in a large amount, .beta.-resorcylic acid is required in the same amount as that of the .alpha.- and .beta.-APM. In the method, the .beta.-resorcylic acid adduct of .alpha.-APM is isolated from a dilute aqueous solution, the .beta.-resorcylic acid is recovered using organic solvents and the like, and the resultant aqueous solution is further concentrated under reduced pressure. Then the isolated .alpha.-APM is recrystallized from water. Separation procedures are, therefore, complex; and the recovery rate of the expensive .alpha.-APM is low, thereby making the method economically unfavorable.
U.S. Pat. No. 3,798,207 discloses a method wherein an only slightly soluble hydrogen halide salt of 60 -APM is formed by bringing .alpha.-APM into contact with hydrogen halide in an aqueous medium in order to separate therefrom .beta.-APM coexisting as an impurity. The method leads to good separation of .alpha.- and .beta.-APM. However, hydrolysis of the methyl ester group of the .alpha.-APM tends to proceed concurrently because a dilute aqueous solution of hydrogen halide is used. Use of the hydrogen halide solution in an excess amount enables good separation of .alpha.-APM from the impurities. On the other hand, a disadvantage the method is that the recovery rate of the hydrogen halide salt of .alpha.-APM is low.
As mentioned above, any separation methods of .alpha.-APM which have been known to date are disadvantageous and unsatisfactory in view of the industrial separation and purification methods involved.